2.1. Methods for Determining Protein Structure
Several algorithms are currently used to evaluate the secondary structure of proteins, including the Kyte-Doolittle, Chou-Fasman-Prevelige, and PHD methods.
The Kyte-Doolittle method (Kyte and Doolittle, 1982, J. Mol. Biol. 157: 105-132) evaluates the hydrophobicity and hydrophilicity of each amino acid, as they appear sequentially in a protein. The program then uses a continuous moving segment approach that determines the average hydropathy within a predetermined segment. Although the program can accurately predict interior and exterior regions of soluble globular proteins, data on membrane spanning regions of transmembrane proteins is more ambiguous.
The Chou-Fasman-Prevelige (CFP) algorithm (Prevelige and Fasman, 1989, in "Predictions of Protein Structure and the Principles of Protein Conformation", Fasman, ed., Plenum Press, New York, pp. 391-416) uses a statistical approach to the study of protein secondary structure. The conformational parameters for each amino acid are calculated using the relative frequency of a given amino acid within a protein, its occurence in a given type of secondary structure, and the fraction of residues occuring in that type of structure. Since these parameters (such as hydrophobicity) contain information about protein stability, properly weighted for their relative importance, they are useful for predicting secondary structures. These parameters, represented by P.alpha. and P.beta. or Pc (for .alpha.-helix, .beta.-sheets or coils, respectively) are utilized to locate nucleation sites within an amino acid sequence. These nucleation sites are then extended until a stretch unlikely to belong to that structure is encountered, whereupon that structure is terminated. This process is repeated throughout the sequence until the secondary structure of the entire sequence is predicted.
The PHD method (Rost and Sander, 1992, Nature 360: 540) utilizes a combination of evolutionary and multiple sequence alignment information, and a "jury" of 12 networks. Since this method is a fully automated computer program, it is independent of human input or interpretation and as such delivers a unique approach.